The enzyme formate dehydrogenase (enzyme no.[EC1.2.1.2]) is an enzyme which catalyzes the reaction giving rise to carbon dioxide and reduced nicotine adenine dinucleotide (hereinafter referred to as NADH) from a formate and oxidized nicotine adenine dinucleotide (hereinafter referred to as NAD). Formate dehydrogenase is a useful enzyme having advantages that when it is used for coenzyme regeneration in a NADH-dependent enzymatic reaction, an inexpensive formate can be utilized and the byproduct is carbon dioxide which is not accumulated in the system. Furthermore, as an enzyme having small Km values for formate and NAD, this enzyme acts effectively at low substrate concentrations and can be utilized for specific microassay of formate, thus being an industrially useful enzyme.
Formate dehydrogenase is known to exist in higher plants, methanol-utilizing yeasts, bacteria, and the like. The enzyme which has heretofore been purified and characterized includes the enzyme derived from the higher plant pea (Pisum sativum: J. Biochem., vol. 77, 845, 1975) and the enzymes derived from methanol-utilizing yeast species, namely Candida boidinii (Eur. J. Biochem., vol. 62, 151, 1976), Candida methylica (Eur. J. Biochem. vol. 152, 657, 1985), Candida methanolica (FEMS Microbiol. Lett., vol. 48, 139, 1987), Kloeckera sp. (Agric. Biol. Chem., vol. 38, 111, 1974), Pichia pastoris (Agric. Biol. Chem., vol. 47, 2547, 1983), and Lipomyces methanosilviensis (JP-A-60-241887), among others. However, these enzymes invariably have some or other shortcomings to be overcome for industrial application, namely low specific activity, large Km values for formate and NAD, and/or a narrow pH range for activity.
There also are several enzymes which have been purified from bacteria and characterized but each has its own drawback in industrial application. For example, the enzymes derived from Pseudomonas sp. 101 (Eur. J. Biochem., vol. 99, 569, 1979) and Pseudomonas oxalaticus (Eur. J. Biochem., vol. 83, 485, 1978) have comparatively high specific activity but are unstable in the absence of a stabilizer. The enzyme derived from Moraxella sp. (J. Bacteriol., vol. 170, 3189, 1988; JP-A-63-313580) is low in specific activity and has a large Km value for formate. The enzyme derived from Hyphomicrobium sp. (JP-A-2000-78970, reported at 1999 Congress of Japan Society for Bioscience, Biotechnology, and Agrochemistry, Synopsis of Papers Read Before the same Congress, p. 234) is low in specific activity and has a low pH range for activity. Moreover, the enzyme derived from Paracoccus sp. (JP-A-03-61481) has a large Km value for formate.
A further problem is that while the products (e.g. haloalcohols) obtainable by reducing haloketone compounds are very useful compounds as raw materials of such as pharmaceutical products, the formate dehydrogenases heretofore reported are invariably inactivated in the presence of a haloketone compound so that if added to an enzymatic reduction reaction system for a haloketone compound, none are able to regenerate the coenzyme. Thus, there is not known a formate dehydrogenase capable of regenerating the coenzyme with good efficiency in the enzymatic reduction reaction system of a haloketone compound.
Referring to formate dehydrogenases of the bacterial origin, enzymes which do not require NAD as the electron acceptor [EC1.2.2.1]), such as those derived from Escherichia coli (J. Biol. Chem., vol. 250, 6693, 1975), Clostridium pasteurianum (J. Bacteriol., vol. 159, 375, 1984), Clostridium thermoaceticum (J. Biol. Chem., vol. 259, 1826, 1983), etc. are also known but these cannot be utilized for the purpose of regenerating the coenzyme NAD.
Referring to the expression of bacterial NAD-dependent formate dehydrogenase genes in transformants, the genes derived from Pseudomonas sp. 101 (Biotechnol. Appl. Biochem., vol. 18, 201, 1993), Mycobacterium vaccae (Appl. Microbiol. Biotechnol., vol. 44, 479, 1995, JP-A-10-23896), Pyrococcus KOD1 (JP-A-2000-69971), Hyphomicrobium sp. (JP-A-2000-78970, reported at the 1999 Congress of Japan Society for Bioscience, Biotechnology, and Agrochemistry, Synopsis of Papers Read Before the same Congress, p. 234) but there has been no knowing of a formate dehydrogenase gene derived from any microorganism of the genus Thiobacillus. 
It is known that bacteria of the genus Thiobacillus have formate dehydrogenase activity in the form of crude enzyme solutions (J. Bacteriol., 131, 389 (1977)) but there is no report on the actual purification and isolation of an enzyme or on the characterization thereof, nor has it been reported that a relevant gene was ever isolated.